Equine infectious anemia (EIA), commonly known as "swamp fever," is a disease which primarily affects horses and ponies. The disease is world wide in distribution and has been extensively investigated for many years to elucidate the morphological and physicochemical properties of the virus, to develop a satifactory diagnostic test for the virus, to find a possible vaccine and to study the epideminology and spread of the infection.
One of the major problems associated with equine infectious anemia is that until recently little or no antibody response could be detected. Recently, researchers have described complement-fixing and neutralizing-antibody responses to this virus, as well as a procedure for purification and concentration of the virus, (Nakajima et al., Arch. Gesamte Virusforsch 28: 348 - 360 (1969).
Despite the advances made in connection with describing some of the characteristics of the virus, a development of a satisfactory diagnostic test for EIA has defied the efforts of many of the prior art workers. Practically all conceivable tests and technics have been attempted at one time or another, and until only very recently, no satisfactory means of assaying the virus or antibody has been available. The use of horse-leukocyte cultures in conjunction with the complement-fixation to assay the virus and antibody has been described (Kobayashi et al., Nat. Inst. Anim. Health Quart. 7: 8 - 20 (1967) and Kono et al., Nat. Inst. Anim. Health Quart. 7: 138 - 144 (1967) and the results appear to explain many of the unique features of the disease; however, the procedure has not been confirmed by other prior art workers.
As it is well known by prior art workers studying EIA, acute and chronic cases of EIA can often be recognized by the clinical symptoms exhibited. However, clinically recovered carriers without the aid of a good diagnostic test may escape the suspicion of even the most astute observer. For example, it has been described that chronically infected horses have been observed experimentally to appear healthy for many years and yet persistently maintain virulent EIA virus in their blood stream. As a result of the dilemma of being unable to ascertain whether an animal is chronically infected with EIA, disease control officials have been in the predicament of trying to prevent the spread of EIA. Thus, the detection of a carrier horse has required the use of the animal inoculation test which usually takes about 90 days and costs about $500.00.
The immunodiffusion reaction technique has been a useful diagnostic and analytical tool for several known diseases. The immunodiffusion reaction involves conducting a reaction between an antigen and an antibody. This reaction is generally effected in a semisolid media such as agar. Both reactants are initially soluble in the medium or carrier, i.e., they diffuse or permeate through the matrix material, but the reaction product (or complex) is insoluble therein and may be observed visually or photographically. A more detailed discussion regarding the immunodiffusion reaction may be found in "Immunodiffusion", by A. J. Crowle, Academic Press, New York, the disclosure of which is incorporated herein by reference.
The success of employing the immunodiffusion reaction technique as a diagnostic test is often dependent upon a relatively pure antigen, which is not substantially tied up or contaminated with antibodies to the antigen. Also, the presence of other contaminating materials will hamper the success of this technique. The titer of the antigen, therefore, must be relatively high in the immunodiffusion reaction technique in order to form a precipitin line of identity with the antiserum. Likewise, the titer of the antiserum must be sufficient so that the antigen and antiserum will form a precipitate. Until recently, antigen for EIA having a high enough titer to form a precipitin line with antiserum has not been available, and, accordingly the use of the immunodiffusion reaction has not been considered by the prior art workers an available tool in diagnosing EIA.